Hemostasis is the spontaneous process of stopping bleeding from damaged blood vessels. Precapillary vessels contract immediately when cut; within seconds, thrombocytes, or blood platelets, are bound to the exposed matrix of the injured vessel by a process called platelet adhesion. Platelets also stick to each other in a phenomenon known as platelet aggregation to form a platelet plug to stop bleeding quickly.
An intravascular thrombus results from a pathological disturbance of hemostasis. Platelet adhesion and aggregation are critical events in intravascular thrombosis. Activated under conditions of turbulent blood flow in diseased vessels or by the release of mediators from other circulating cells and damaged endothelial cells lining the vessel, platelets accumulate at a site of vessel injury and recruit further platelets into the developing thrombus. The thrombus can grow to sufficient size to block off arterial blood vessels. Thrombi can also form in areas of stasis or slow blood flow in veins. Venous thrombi can easily detach portions of themselves called emboli that travel through the circulatory system and can result in blockade of other vessels, such as pulmonary arteries. Thus, arterial thrombi cause serious disease by local blockade, whereas venous thrombi do so primarily by distant blockade, or embolization. These conditions include venous thrombosis, thrombophlebitis, arterial embolism, coronary and cerebral arterial thrombosis, unstable angina, myocardial infarction, stroke, cerebral embolism, kidney embolisms and pulmonary embolisms.
A number of converging pathways lead to platelet aggregation. Whatever the initial stimulus, the final common event is crosslinking of platelets by binding fibrinogen to a membrane binding site, glycoprotein IIb/IIIa (GPIIb/IIIa). Compounds that are antagonists for GPIIb/IIIa receptor complex have been shown to inhibit platelet aggregation (U.S. Pat. Nos. 6,037,343 and 6,040,317). Antibodies against GPIIb/IIIa have also been shown to have high antiplatelet efficacy (The EPIC investigators, New Engl. J. Med. (1994) 330:956-961). However, this class of antiplatelet agents sometimes causes bleeding problems.
Thrombin can produce platelet aggregation largely independently of other pathways but substantial quantities of thrombin are unlikely to be present without prior activation of platelets by other mechanisms. Thrombin inhibitors such as hirudin are highly effective antithrombotic agents. However, functioning as both antiplatelet and anti-coagulant agents, thrombin inhibitors again can produce excessive bleeding. (The TIMI 9a investigators, The GUSTO Iia investigators, Circulation, 90: 1624-1630 (1994); Circulation, 90: 1631-1637 (1994); Neuhaus K. L. et al., Circulation, 90: 1638-1642 (1994).)
Various antiplatelet agents have been studied for many years as potential targets for inhibiting thrombus formation. Some agents such as aspirin and dipyridamole have come into use as prophylactic antithrombotic agents, and others have been the subjects of clinical investigations. To date, the powerful agents such as disintegrins, and the thienopyridines ticlopidine and clopidogrel have been shown to have substantial side effects, while agents such as aspirin have useful but limited effectiveness (Hass, et al., N. Engl. J. Med., 321:501-507 (1989); Weber, et al., Am. J. Cardiol. 66:1461-1468 (1990); Lekstrom and Bell, Medicine 70:161-177 (1991)). In particular, use of the thienopyridines in antiplatelet therapy has been shown to increase the incidence of potentially life threatening thrombotic thrombocytopenic purpura (Bennett, C. L. et al. N. Engl. J. Med, (2000) 342: 1771-1777). Aspirin, which has a beneficial effect on platelet aggregation (Br. Med. J. (1994) 308: 81-106; 159-168), acts by inducing blockade of prostaglandin synthesis. Aspirin has no effect on ADP-induced platelet aggregation, and thus has limited effectiveness on platelet aggregation. Furthermore, its well-documented high incidence of gastric side effects limits its usefulness in many patients. Clinical efficacy of some newer drugs, such as ReoPro (7E3), is impressive, but recent trials have found that these approaches are associated with an increased risk of major bleeding, sometimes necessitating blood transfusion (New Engl. J. Med. (1994) 330:956-961). Thus it appears that the ideal “benefit/risk” ratio has not been achieved.
Recent studies have suggested that adenosine 5′-diphosphate (ADP), a common agonist, plays a key role in the initiation and progression of arterial thrombus formation (Bernat, et al., Thromb. Haemostas. (1993) 70:812-826); Maffrand, et al., Thromb. Haemostas. (1988) 59:225-230; Herbert, et al., Arterioscl. Thromb. (1993) 13:1171-1179). ADP induces platelet aggregation, shape change, secretion, influx and intracellular mobilization of Ca+2, and inhibition of adenylyl cyclase. Binding of ADP to platelet receptors is required for elicitation of the ADP-induced platelet responses. There are at least three P2 receptors expressed in human platelets: a cation channel receptor P2X1, a G protein-coupled receptor P2Y1, and a G protein-coupled receptor P2Y12 (also referred to as P2Yac and P2T) The P2X1 receptor is responsible for rapid calcium influx and is activated by ATP. The role of P2X1 receptors in the process of platelet aggregation is not fully understood. However, it has been suggested that the P2X1 receptor participates in platelet shape change (Rolf, et al., Thromb Haemost. 85:303-308, 2001), and in platelet thombi formation under high shear forces. (Hechler, et al., J Exp Med. 198: 661-667, 2003). The P2Y1 receptor is responsible for calcium mobilization, shape change and the initiation of aggregation. P2Y12 receptor is responsible for inhibition of adenylyl cyclase and is required for full aggregation. (Hourani, et al., The Platelet ADP Receptors Meeting, La Thuile, Italy, Mar. 29-31, 2000.)
Ingall et al. (J. Med. Chem. 42: 213-220, (1999)) describe a dose-related inhibition of ADP-induced platelet aggregation by analogues of adenosine triphosphate (ATP), which is a weak, nonselective but competitive P2Y12 receptor antagonist. Zamecnik (U.S. Pat. No. 5,049,550) discloses a method for inhibiting platelet aggregation in a mammal by administering to said mammal a diadenosine tetraphosphate compound of App(CH2)ppA or its analogs. Kim et al. (U.S. Pat. No. 5,681,823) disclose P1, P4-dithio-P2, P3-monochloromethylene 5′, 5′″ diadenosine P1, P4-tetraphosphate as an antithrombotic agent. The thienopyridines ticlopidine and clopidogrel, which are metabolized to antagonists of the platelet P2Y12 receptor, are shown to inhibit platelet function in vivo (Quinn and Fitzgerald, Circulation 100:1667-1672 (1999); Geiger, et al., Arterioscler. Thromb. Vasc. Biol. 19:2007-2011 (1999)). However, these thienopyridines have a number of therapeutic disadvantages:                Slow onset of action (Gurbel, et al., Am J. Cardiol. 90: 312-315, 2002)        Due to the irreversible nature of these inhibitors on the P2Y12 receptor, subjects treated with thienopyridines are at a high risk of bleeding if a surgical procedure is necessary. For elective surgeries, discontinuation of the drug is necessary for at least five to ten days since production of new platelets is necessary to restore hemostasis, exposing the subject to a high risk of thrombotic events during this period (Kapetanakis, et al., Eur Heart J. 26: 576-583, 2005).        Subjects treated with the standard dose regimen of these compounds present a large inter-individual variability in the pharmacological effect of the drug, with a significant proportion of patients underprotected from the occurrence of ischemic events. (Gurbel, et al., Circulation 107: 2908-2913, 2003; Aleil, et al., J Thromb Haemost. 3: 85-92, 2005).        
Stents are typically slotted metal tubes, which can be expanded by a balloon in an angioplastied artery, providing a rigid structural support for the arterial wall. The use of coronary stents for the treatment of patients with acute coronary syndrome has increased significantly during the past years. With coronary stents implanted in more than 2 million people worldwide, some doctors and researchers are now concerned about a long-term problem of blood clots inside the stents that is observed in some patients who have received stents.
In-stent restenosis is caused primarily due to hyperplasia of smooth muscle cells in the intimal layer of the vessel wall (so-called neointimal hyperplasia) and, to a much lesser extent, mural thrombus. On the molecular and cellular levels, the initial vascular injury caused by both inflation of intracoronary balloons and the metal of the stent itself results in denudation of the intima and stretching of the media and adventitia, in addition, both macrophages and polymorphonuclear neutrophils migrate to the site of damage, where they release chemokines. These chemokines serve to increase the amount of matrix metalloproteinase, which leads to remodeling of the extracellular matrix and stimulate smooth muscle cell migration. The wound healing reaction stimulates platelets, growth factor and smooth muscle cell activation, followed by smooth muscle cell and fibroblast migration and proliferation into the injured area. Smooth muscle cells are also stimulated to increase the expression of genes involved in cell division. It is both the interaction and the extent of these processes that lead to neointimal hyperplasia and in-stent restenosis, which are characterized by a marked proliferative response produced by the stent as has been demonstrated by histological examinations. Stenting also raises the systemic levels of inflammatory markers such as C-reactive protein and interleukin-6.
Recently, stents are coated with agents that reduce or prevent exaggerated neointimal proliferation, and thereby, restenosis. For example, paclitaxel-eluting stents inhibit the proliferation of smooth muscle cells, and sirolimus-eluting stents inhibits the inflammation response of the arterial wall. One problem with these stents is that the drugs also inhibit the regeneration of the endothelium destroyed during the expansion of the narrowed artery, creating a potential risk of thrombosis. Thus, the placement of these stents often requires the treatment by systemic administration of antithrombotic drugs.
There is a need in the area of cardiovascular and cerebrovascular therapeutics for improved stents.